[摘要] 目的：探讨lncRNA POU3F3 通过调节MGMT的表达影响高级别脑胶质瘤细胞对替莫唑胺(temozolomide，TMZ)耐药及其作用机制。方法：选取2016 年1 月至2018 年1 月北京大学国际医院神经外科收治的60 例患者组织标本，其中，脑外伤患者12 例（正常组）、初发高级别脑胶质瘤患者30 例（初发组）、复发高级别脑胶质瘤患者[复发组，已经接受了手术治疗+TMZ综合治疗再次复发的人群]18 例。采用1、2、4 及8 μg/ml TMZ诱导U251 细胞并维持正常生长1 周以构建抵抗U251 TMZ耐药细胞系（U251 TMZ-resistance，U251-TR）；正常组采用相同体积的0.9%生理盐水处理U251 细胞。采用qPCR和Wb检测正常脑组织及神经胶质瘤细胞中POU3F3 和甲基鸟嘌呤-DNA甲基转移酶（methylguanine DNA methyltransferase，MGMT）mRNA和蛋白的表达水平。慢病毒转染构建稳定干扰POU3F3 的U251-TR细胞系（U251-TR stable interference POU3F3，U251-TR siPOU3F3），MTT法验证U251-TR 细胞并检测细胞TMZ IC50值，Wb检测细胞中MGMT蛋白的表达水平。结果：与正常组及初发组比较，复发组POU3F3 表达显著增高（P<0.01），U251-TR细胞TMZ IC50值显著高于U251 细胞（P<0.01），U251-TR siPOU3F3 细胞TMZ IC50值显著低于U251-TR细胞（均P<0.01），但高于U251 细胞（P<0.01）；U251-TR细胞POU3F3 及MGMT mRNA和蛋白的表达水平均高于U251 细胞（均P<0.01），U251-TR siPOU3F3 细胞POU3F3 及MGMT mRNA和蛋白表达水平均低于U251-TR 细胞（均P<0.01）。结论：lncRNA POU3F3 是促进高级别脑胶质瘤细胞TMZ耐药性的关键因素，可能对临床上TMZ耐药的研究有一定的意义。

[Abstract] Objective: To explore the mechanism of long non-coding RNA POU3F3 (lncRNAPOU3F3) affecting temozolomide (TMZ)-resistance in high-grade glioma cells via regulating MGMT expression. Methods: Sixty cases of tissues from patients treated at the Department of Neurosurgery, Peking University International Hospital during January 2016 and January 2018 were collected for this study, including 12 cases from brain trauma patients (normal group), 30 cases from primary high-grade glioma patients (primary onset group) and 18 cases from recurrent high-grade glioma patients (recurrence group, accepted surgery+TMZ already). U251 cells were induced with TMZ at the concentration of 1, 2, 4 and 8 μg/ml and maintained normal growth for a week to construct TMZ-resistant U251cell line (U251 TMZ-resistance, U251-TR); and the normal control group was treated with equal volume of physiological saline.Reverse transcription polymerase chain reaction (qPCR) and Wb were used to detect the mRNA and protein expressions of POU3F3 and MGMT (methylguanine DNA methyltransferase) in normal brain tissues and glioma cells. Lentivirus transfection was used to construct U251 cell line with stable POU3F3 interference (U251-TR siPOU3F3); CCK-8 was used to detect TMZ IC50 value (the half maximal inhibitory concentration) in each group of U251 cells, and Wb was used to detect the expression of MGMT protein in each group of cells. Results: Compared with the normal group and primaryonset group, the expression of POU3F3 in recurrence group was significantly increased (P<0.01). The TMZ IC50 of U251-TR cells was significantly higher than that of U251 cells (P<0.01), and The TMZ IC50 of U251-TR siPOU3F3 cells was significantly lower than that of U251-TR cellsbut higher than that of U251 cells (all P<0.01). The protein and mRNA expressions of POU3F3 and MGMT in U251-TR cells were significantly higher than that in U251 cells (P<0.01),while those expressions in U251-TR siPOU3F3 cells were significantly lower than those in U251-TR cells (P<0.01).Conclusion: lncRNA POU3F3 is the key factor to promote TMZ resistance in human high-grade gliomas cells, which may exert certain guiding significance in the clinical treatment for TMZ resistance.

首都卫生发展科研专项基金资助项目（No.2018-2-1072）